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Cloning and identification of a novel plasma membrane intrinsic protein gene ScPIP2-1 in sugarcane

汤翰臣  Qing Yu  Zhu Li  Feng Liu  Weihua Su  Hui Ling  Yachun Su  Youxiong Que  
【摘要】:【Research background】 Sugarcane is the most important sugar crop and a major source of sugar production in the world. In our country, 80% of the sugarcane planting areas are dry sloping fields, which are characterized by low rainfall, uneven rainfall and low soil fertility. They are prone to drought and high salt stress, resulting in the loss of total yield and quality of sugarcane. Aquaporins(AQPs) play an important role in maintaining water balance, regulating plant growth and resisting stress, which can be found in major intrinsic protein(MIP) super family and has explicit classification, including plasma membrane intrinsic proteins(PIPs), tonoplast membrane intrinsic proteins(TIPs), nodulin26-like major intrinsic proteins(NIPs), small and basic intrinsic proteins(SIPs) and X intrinsic proteins(XIPs). PIP2 s, one of the family members of PIPs, was found to be responded to osmotic stress such as water balance, water deficit and salinity. So far, PIP2 s have not been reported in sugarcane. 【Materials and methods】 A full-length cDNA sequence of Sc PIP2-1 was successfully cloned from the Saccharum spp. hybrid(ROC22) by RT-PCR. Preliminary identification of ScPIP2-1 gene function was confirmed by tissue-specific expression analysis, gene expression patterns analysis of ScPIP2-1 under polyethylene glycol(PEG), salt(NaCl) and abscisic acid(ABA) stresses, subcellular localization, and transcriptional activation activity analysis by yeast two hybrid self-activation experiment, gene transient expression analysis under drought and NaCl in Nicotiana benthamiana leaves. 【Result and analysis】 The cDNA length of ScPIP2-1 gene was 1145 bp, containing a 873 bp complete open reading frame and encoding 290 amino acids residues. qRT-PCR analysis showed that ScPIP2-1 was constitutively expressed in all five collected tissues(root, leaf, bud, flesh and peel), mainly accumulated in leaves, roots and buds, and with the highest in buds but the lowest in peels. The expression levels of Sc PIP2-1 gene in both ROC22 underground and aboveground tissues were all up regulated under ABA, PEG and NaCl stresses. Confocal microscopy study indicated that ScPIP2-1 was located in cytomembrane and cytoplasm. The result of yeast hybridization experiment demonstrated that ScPIP2-1 did not possess transcriptional activity. The results from chlorophyll meter and conductivity meter showed that ScPIP2-1 responsed earlier to drought than NaCl stress. 【Conclusion】 These results suggest that sugarcane ScPIP2-1 gene may play a positive role in the response to drought and salt stresses.

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